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July 7, 2019  |  

MethSMRT: an integrative database for DNA N6-methyladenine and N4-methylcytosine generated by single-molecular real-time sequencing

DNA methylation is an important type of epigenetic modifications, where 5- methylcytosine (5mC), 6-methyadenine (6mA) and 4-methylcytosine (4mC) are the most common types. Previous efforts have been largely focused on 5mC, providing invaluable insights into epigenetic regulation through DNA methylation. Recently developed single-molecule real-time (SMRT) sequencing technology provides a unique opportunity to detect the less studied DNA 6mA and 4mC modifications at single-nucleotide resolution. With a rapidly increased amount of SMRT sequencing data generated, there is an emerging demand to systematically explore DNA 6mA and 4mC modifications from these data sets. MethSMRT is the first resource hosting DNA 6mA and 4mC methylomes. All the data sets were processed using the same analysis pipeline with the same quality control. The current version of the database provides a platform to store, browse, search and download epigenome-wide methylation profiles of 156 species, including seven eukaryotes such as Arabidopsis, C. elegans, Drosophila, mouse and yeast, as well as 149 prokaryotes. It also offers a genome browser to visualize the methylation sites and related information such as single nucleotide polymorphisms (SNP) and genomic annotation. Furthermore, the database provides a quick summary of statistics of methylome of 6mA and 4mC and predicted methylation motifs for each species. MethSMRT is publicly available at http://sysbio.sysu.edu.cn/methsmrt/ without use restriction.

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July 7, 2019  |  

Complete genome sequences of four different Bordetella sp. isolates causing human respiratory infections.

Species of the genus Bordetella associate with various animal hosts, frequently causing respiratory disease. Bordetella pertussis is the primary agent of whooping cough and other Bordetella species can cause similar cough illness. Here, we report four complete genome sequences from isolates of different Bordetella species recovered from human respiratory infections. Copyright © 2016 Weigand et al.

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July 7, 2019  |  

Draft genome sequence of the environmentally isolated Acinetobacter pittii strain IPK_TSA6.1.

Acinetobacter pittii is an opportunistic pathogen frequently isolated from Acinetobacter infections other than those from Acinetobacter baumannii Multidrug resistance in A. pittii, including resistance to carbapenems, has been increasingly reported worldwide. Here, we report the 4.14-Mbp draft genome sequence of A. pittii IPK_TSA6.1 that was isolated from a nonhospital setting. Copyright © 2016 Lee and Jang.

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July 7, 2019  |  

Genetic characterization of a blaVEB-2-carrying plasmid in Vibrio parahaemolyticus.

This study reports the first detection of blaVEB-2 gene in Vibrio parahaemolyticus strain isolated from a shrimp sample. The blaVEB-2 was carried on a novel Inc type plasmid, was likely to originate from aquatic organisms upon comparison with other known genetic elements in the GenBank. However, the plasmid contains resistance elements usually harbored by members of Enterobacteriaceae, suggesting that gene transfer events occurred and contributed to the formation of this multidrug resistance-encoding plasmid. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

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July 7, 2019  |  

Use of WGS data for investigation of a long-term NDM-1-producing Citrobacter freundii outbreak and secondary in vivo spread of blaNDM-1 to Escherichia coli, Klebsiella pneumoniae and Klebsiella oxytoca.

An outbreak of NDM-1-producing Citrobacter freundii and possible secondary in vivo spread of blaNDM-1 to other Enterobacteriaceae were investigated.From October 2012 to March 2015, meropenem-resistant Enterobacteriaceae were detected in 45 samples from seven patients at Aalborg University Hospital, Aalborg, Denmark. In silico resistance genes, Inc plasmid types and STs (MLST) were obtained from WGS data from 24 meropenem-resistant isolates (13 C. freundii, 6 Klebsiella pneumoniae, 4 Escherichia coli and 1 Klebsiella oxytoca) and 1 meropenem-susceptible K. oxytoca. The sequences of the meropenem-resistant C. freundii isolates were compared by phylogenetic analyses. In vitro susceptibility to 21 antimicrobial agents was tested. Furthermore, in vitro conjugation and plasmid characterization was performed.From the seven patients, 13 highly clonal ST18 NDM-1-producing C. freundii were isolated. The ST18 NDM-1-producing C. freundii isolates were only susceptible to tetracycline, tigecycline, colistin and fosfomycin (except for the C. freundii isolates from Patient 2 and Patient 7, which were additionally resistant to tetracycline). The E. coli and K. pneumoniae from different patients belonged to different STs, indicating in vivo transfer of blaNDM-1 in the individual patients. This was further supported by in vitro conjugation and detection of a 154 kb IncA/C2 plasmid with blaNDM-1. Patient screenings failed to reveal any additional cases. None of the patients had a history of recent travel abroad and the source of the blaNDM-1 plasmid was unknown.To our knowledge, this is the first report of an NDM-1-producing C. freundii outbreak and secondary in vivo spread of an IncA/C2 plasmid with blaNDM-1 to other Enterobacteriaceae.© The Author 2016. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

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July 7, 2019  |  

Full-length nucleotide sequences of mcr-1-harboring plasmids isolated from extended- spectrum-ß-lactamase-producing Escherichia coli isolates of different origins.

Here, we present the full sequences of three mcr-1-carrying plasmids isolated from extended-spectrum-ß-lactamase (ESBL)-producing Escherichia coli The plasmids belong to three different replicon types and are 34,640 bp, 209,401 bp, and 247,885 bp in size. We describe for the first time a composite transposon containing mcr-1 localized on a multidrug-resistant (MDR) IncHI2 plasmid harboring additional determinants of resistance to six different classes of antibiotics, including the ESBL gene blaCTX-M-1, and heavy metal resistance. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

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July 7, 2019  |  

Whole genome sequence analysis indicates recent diversification of mammal-associated Campylobacter fetus and implicates a genetic factor associated with H2S production.

Campylobacter fetus (C. fetus) can cause disease in both humans and animals. C. fetus has been divided into three subspecies: C. fetus subsp. fetus (Cff), C. fetus subsp. venerealis (Cfv) and C. fetus subsp. testudinum (Cft). Subspecies identification of mammal-associated C. fetus strains is crucial in the control of Bovine Genital Campylobacteriosis (BGC), a syndrome associated with Cfv. The prescribed methods for subspecies identification of the Cff and Cfv isolates are: tolerance to 1 % glycine and H2S production.In this study, we observed the deletion of a putative cysteine transporter in the Cfv strains, which are not able to produce H2S from L-cysteine. Phylogenetic reconstruction of the core genome single nucleotide polymorphisms (SNPs) within Cff and Cfv strains divided these strains into five different clades and showed that the Cfv clade and a Cff clade evolved from a single Cff ancestor.Multiple C. fetus clades were observed, which were not consistent with the biochemical differentiation of the strains. This suggests the need for a closer evaluation of the current C. fetus subspecies differentiation, considering that the phenotypic differentiation is still applied in BGC control programs.

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July 7, 2019  |  

Tracking inter-institutional spread of NDM and identification of a novel NDM-positive plasmid, pSg1-NDM, using next-generation sequencing approaches.

Owing to gene transposition and plasmid conjugation, New Delhi metallo-ß-lactamase (NDM) is typically identified among varied Enterobacteriaceae species and STs. We used WGS to characterize the chromosomal and plasmid molecular epidemiology of NDM transmission involving four institutions in Singapore.Thirty-three Enterobacteriaceae isolates (collection years 2010-14) were sequenced using short-read sequencing-by-synthesis and analysed. Long-read single molecule, real-time sequencing (SMRTS) was used to characterize genetically a novel plasmid pSg1-NDM carried on Klebsiella pneumoniae ST147.In 20 (61%) isolates, blaNDM was located on the pNDM-ECS01 plasmid in the background of multiple bacterial STs, including eight K. pneumoniae STs and five Escherichia coli STs. In six (18%) isolates, a novel blaNDM-positive plasmid, pSg1-NDM, was found only in K. pneumoniae ST147. The pSg1-NDM-K. pneumoniae ST147 clone (Sg1-NDM) was fully sequenced using SMRTS. pSg1-NDM, a 90?103 bp IncR plasmid, carried genes responsible for resistance to six classes of antimicrobials. A large portion of pSg1-NDM had no significant homology to any known plasmids in GenBank. pSg1-NDM had no conjugative transfer region. Combined chromosomal-plasmid phylogenetic analysis revealed five clusters of clonal bacterial NDM-positive plasmid transmission, of which two were inter-institution clusters. The largest inter-institution cluster involved six K. pneumoniae ST147-pSg1-NDM isolates. Fifteen patients were involved in transmission clusters, of which four had ward contact, six had hospital contact and five had an unknown transmission link.A combined sequencing-by-synthesis and SMRTS approach can determine effectively the transmission clusters of blaNDM and genetically characterize novel plasmids. Plasmid molecular epidemiology is important to understanding NDM spread as blaNDM-positive plasmids can conjugate extensively across species and STs.© The Author 2016. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

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July 7, 2019  |  

Complete sequencing of plasmids containing blaOXA-163 and blaOXA-48 in Escherichia coli ST131.

OXA-48-like enzymes have emerged as important extended-spectrum ß-lactamases/carbapenemases in E. coli ST131. We report the structure of the first fully sequenced blaOXA-163 plasmid, and of two other blaOXA-48 plasmids in this lineage. blaOXA-163 was located on a 71kb IncN plasmid with other resistance genes. blaOXA-48 was present on IncL/M plasmids, genetically similar to other blaOXA-48 plasmid sequences, and consistent with inter-species/inter-lineage spread. The presence of blaOXA-48-like genes on epidemic plasmids in ST131 is of concern. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

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July 7, 2019  |  

Plasmids from Shiga toxin-producing Escherichia coli strains with rare enterohemolysin gene (ehxA) subtypes reveal pathogenicity potential and display a novel evolutionary path.

Most Shiga toxin-producing Escherichia coli (STEC) strains associated with severe disease, such as hemolytic-uremic syndrome (HUS), carry large enterohemolysin-encoding (ehxA) plasmids, e.g., pO157 and pO103, that contribute to STEC clinical manifestations. Six ehxA subtypes (A through F) exist that phylogenetically cluster into eae-positive (B, C, F), a mix of eae-positive (E) and eae-negative (A), and a third, more distantly related, cluster of eae-negative (D) STEC strains. While subtype B, C, and F plasmids share a number of virulence traits that are distinct from those of subtype A, sequence data have not been available for subtype D and E plasmids. Here, we determined and compared the genetic composition of four subtype D and two subtype E plasmids to establish their evolutionary relatedness among ehxA subtypes and define their potential role in pathogenicity. We found that subtype D strains carry one exceptionally large plasmid (>200 kbp) that carries a variety of virulence genes that are associated with enterotoxigenic and enterohemorrhagic E. coli, which, quite possibly, enables these strains to cause disease despite being food isolates. Our data offer further support for the hypothesis that this subtype D plasmid represents a novel virulence plasmid, sharing very few genetic features with other plasmids; we conclude that these plasmids have evolved from a different evolutionary lineage than the plasmids carrying the other ehxA subtypes. In contrast, the 50-kbp plasmids of subtype E (pO145), although isolated from HUS outbreak strains, carried only few virulence-associated determinants, suggesting that the clinical presentation of subtype E strains is largely a result of chromosomally encoded virulence factors.Bacterial plasmids are known to be key agents of change in microbial populations, promoting the dissemination of various traits, such as drug resistance and virulence. This study determined the genetic makeup of virulence plasmids from rare enterohemolysin subtype D and E Shiga toxin-producing E. coli strains. We demonstrated that ehxA subtype D plasmids represent a novel E. coli virulence plasmid, and although subtype D plasmids were derived from nonclinical isolates, they encoded a variety of virulence determinants that are associated with pathogenic E. coli In contrast, subtype E plasmids, isolated from strains recovered from severely ill patients, carry only a few virulence determinants. The results of this study reemphasize the plasticity and vast diversity among E. coli plasmids. This work demonstrates that, although E. coli strains of certain serogroups may not be frequently associated with disease, they should not be underestimated in protecting human health and food safety. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

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July 7, 2019  |  

Genomic recombination leading to decreased virulence of group B Streptococcus in a mouse model of adult invasive disease.

Adult invasive disease caused by Group B Streptococcus (GBS) is increasing worldwide. Whole-genome sequencing (WGS) now permits rapid identification of recombination events, a phenomenon that occurs frequently in GBS. Using WGS, we described that strain NGBS375, a capsular serotype V GBS isolate of sequence type (ST)297, has an ST1 genomic background but has acquired approximately 300 kbp of genetic material likely from an ST17 strain. Here, we examined the virulence of this strain in an in vivo model of GBS adult invasive infection. The mosaic ST297 strain showed intermediate virulence, causing significantly less systemic infection and reduced mortality than a more virulent, serotype V ST1 isolate. Bacteremia induced by the ST297 strain was similar to that induced by a serotype III ST17 strain, which was the least virulent under the conditions tested. Yet, under normalized bacteremia levels, the in vivo intrinsic capacity to induce the production of pro-inflammatory cytokines was similar between the ST297 strain and the virulent ST1 strain. Thus, the diminished virulence of the mosaic strain may be due to reduced capacity to disseminate or multiply in blood during a systemic infection which could be mediated by regulatory factors contained in the recombined region.

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July 7, 2019  |  

Comparative genomics of Campylobacter iguaniorum to unravel genetic regions associated with reptilian hosts.

Campylobacter iguaniorum is most closely related to the species C fetus, C hyointestinalis, and C lanienae Reptiles, chelonians and lizards in particular, appear to be a primary reservoir of this Campylobacter species. Here we report the genome comparison of C iguaniorum strain 1485E, isolated from a bearded dragon (Pogona vitticeps), and strain 2463D, isolated from a green iguana (Iguana iguana), with the genomes of closely related taxa, in particular with reptile-associated C fetus subsp. testudinum In contrast to C fetus, C iguaniorum is lacking an S-layer encoding region. Furthermore, a defined lipooligosaccharide biosynthesis locus, encoding multiple glycosyltransferases and bounded by waa genes, is absent from C iguaniorum Instead, multiple predicted glycosylation regions were identified in C iguaniorum One of these regions is > 50 kb with deviant G + C content, suggesting acquisition via lateral transfer. These similar, but non-homologous glycosylation regions were located at the same position on the genome in both strains. Multiple genes encoding respiratory enzymes not identified to date within the C. fetus clade were present. C iguaniorum shared highest homology with C hyointestinalis and C fetus. As in reptile-associated C fetus subsp. testudinum, a putative tricarballylate catabolism locus was identified. However, despite colonizing a shared host, no recent recombination between both taxa was detected. This genomic study provides a better understanding of host adaptation, virulence, phylogeny, and evolution of C iguaniorum and related Campylobacter taxa. © The Author 2016. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.

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